12 blot of B. mallei lysate (lanes 2 and 3) and OMV (lanes 4 and 5) preparations using pooled sera from. 13 sham-immunized (lane 2) or OMV immunized (lanes ...
4
A Burkholderia pseudomallei Outer Membrane Vesicle Vaccine Provides Cross Protection against Inhalational Glanders in Mice and Non-Human Primates
5 6
Sarah M. Baker, Christopher J. H. Davitt, Natalya Motyka, Nicole L. Kikendall, Kasi RussellLodrigue, Chad J. Roy, and Lisa A. Morici
1 2 3
7 8 9 10
Figure S1. OMVs purified from B. pseudomallei strain Bp82. Purified OMVs were negatively stained with 1% uranyl acetate and imaged by transmission electron microscopy. OMVs range between 25 and 200 nm in diameter. Magnification 31,000x. Scale bar 200nm.
Vaccines 2017, 5, 49; doi:10.3390/vaccines5040049
www.mdpi.com/journal/vaccines
Vaccines 2017, 5, 49
2 of 3
Control
B.mallei
OMV
OMV
2
3
4
5
1 kD 250 150 100 75
50 37
25 20 15 10
11 12 13 14 15 16 17
Figure S2. OMV immunization induces IgG antibodies against numerous B. mallei antigens. Western blot of B. mallei lysate (lanes 2 and 3) and OMV (lanes 4 and 5) preparations using pooled sera from sham-immunized (lane 2) or OMV immunized (lanes 3 and 4) mice (n=5 per group). OMV Opolysaccharide (OPS) in lane 5 was detected using a monoclonal antibody (Pp-Ps-W) to B. pseudomallei OPS to facilitate comparison of immunoreactive proteins versus OPS in B. mallei lysate probed with OMV immune sera. Lane 1 = molecular weight marker.
Inhaled CFU
4×106 3×106 2×106 1×106 0
KN25 KR40 KP06 KL22 KM81 KI62 KH26 KL67 KP63 KT28
Sham immunized
OMV immunized
(a) ns
mean inhaled dose
3000000
2000000
1000000
0
Sham
OMV
(b)
18 19 20 21 22
Figure S3. B. mallei aerosol challenge doses for rhesus macaques immunized with sham or OMV vaccine. (a) Inhaled doses of B. mallei were calculated for each macaque. Doses ranged from 7.1x105 to 2.9 x 106 cfu per animal. (b) Mean inhaled dose per immunization group. There was no difference in inhaled dose between sham immunized and OMV-immunized animals (ns = not significant, p=0.77 using students t-test).
Vaccines 2017, 5, 49
3 of 3
800
30
ns
ns 20
relative D (f)
relative D (Mv)
600
400
200
0
-200
10
0
-10
Sham
OMV
(a) 30
-20
OMV
Sham
(b)
ns
relative D (EF50)
20
10
0
-10
-20
Sham
OMV
(c)
23 24 25 26 27 28
Figure S4. Changes in respiratory function after immunization and B. mallei challenge in nonhuman primates. Pulmonary function of rhesus macaques vaccinated with OMVs or saline (shamvaccinated). Relative change (△) in each respiratory parameter prior to challenge when compared to measurement +7days after B. mallei aerosol challenge. (a) Minute volume (ml/minute); (b) frequency (breaths/min); (c) EF50 (ml/sec). Group comparison by Kolmogorov-Smirnov test, ns= not significant at p
